Three Main Goals of this File
Produce Cleaner looking code.
Identify the amount of clusters there are
Identify the top genes
expressed in each of the clusters
##
## The downloaded binary packages are in
## /var/folders/7n/x74qctp91rng390gx0z9hmd80000gn/T//RtmpIXQzec/downloaded_packages
options(future.globals.maxSize = 74 * 1024^3) # 55 GB
getOption("future.globals.maxSize") #59055800320## [1] 79456894976
SO4@meta.data$sample <- factor(
SO4@meta.data$sample,
levels = c("SO1","SO4","SO2","SO3"))
DimPlot(SO4,split.by ="sample",group.by = "sample")markers.to.plot1 <- c(
"Atf3", #
"Egr1", #
"Fos", #
"Jun", #
"Junb", #
"Pappa2", #
"Cxcl10", #
"Cldn19", #
"Krt7", #
"Egf", #
"Ptger3",
"Ckb",
"Mcub",
"Fabp3",
"Foxq1",
"Vash2",
"Pamr1",
"Vegfa"
)
DotPlot(SO4,
features = markers.to.plot1,
dot.scale = 8,
dot.min = 0,
scale = FALSE,
scale.max = 100,
scale.min = 0,
col.min = -2.5,
group.by = "seurat_clusters",
col.max = 2.5)+
coord_flip()##
## Attaching package: 'plotly'
## The following object is masked from 'package:AnnotationDbi':
##
## select
## The following object is masked from 'package:IRanges':
##
## slice
## The following object is masked from 'package:S4Vectors':
##
## rename
## The following object is masked from 'package:ggplot2':
##
## last_plot
## The following object is masked from 'package:stats':
##
## filter
## The following object is masked from 'package:graphics':
##
## layout
Idents(SO4) <- "subclass2_MD"
MD_DEGs <- FindAllMarkers(SO4,
only.pos = TRUE,
logfc.threshold = 0.1,
min.pct = 0.1,
min.diff.pct = 0.1,
return.thresh = 0.05)## Calculating cluster type_1
## Warning: The `slot` argument of `GetAssayData()` is deprecated as of SeuratObject 5.0.0.
## ℹ Please use the `layer` argument instead.
## ℹ The deprecated feature was likely used in the Seurat package.
## Please report the issue at <https://github.com/satijalab/seurat/issues>.
## This warning is displayed once every 8 hours.
## Call `lifecycle::last_lifecycle_warnings()` to see where this warning was
## generated.
## Warning: `PackageCheck()` was deprecated in SeuratObject 5.0.0.
## ℹ Please use `rlang::check_installed()` instead.
## ℹ The deprecated feature was likely used in the Seurat package.
## Please report the issue at <https://github.com/satijalab/seurat/issues>.
## This warning is displayed once every 8 hours.
## Call `lifecycle::last_lifecycle_warnings()` to see where this warning was
## generated.
## For a (much!) faster implementation of the Wilcoxon Rank Sum Test,
## (default method for FindMarkers) please install the presto package
## --------------------------------------------
## install.packages('devtools')
## devtools::install_github('immunogenomics/presto')
## --------------------------------------------
## After installation of presto, Seurat will automatically use the more
## efficient implementation (no further action necessary).
## This message will be shown once per session
## Calculating cluster type_2
## Calculating cluster type_3
## Calculating cluster type_4
MD_DEGs <- MD_DEGs %>% arrange(desc(avg_log2FC))
write.xlsx(MD_DEGs, file = here("jk_code", "DEGs_MD_SUBclass.xlsx"), rowNames = TRUE)
# save as excel file
MD_DEGs %>%
group_by(cluster) %>%
dplyr::filter(avg_log2FC > 1, p_val_adj < 0.05) %>%
slice_head(n = 10) %>%
ungroup() -> top10
# save as excel file
DoHeatmap(SO4, features = top10$gene) + NoLegend()## Warning in DoHeatmap(SO4, features = top10$gene): The following features were
## omitted as they were not found in the scale.data slot for the SCT assay: Oasl2
Idents(SO4) <- "subclass_MD"
MD_DEGs2 <- FindAllMarkers(SO4,
only.pos = TRUE,
logfc.threshold = 0.1,
min.pct = 0.1,
min.diff.pct = 0.1,
return.thresh = 0.05)## Calculating cluster type_1
## Calculating cluster type_2a
## Calculating cluster type_2b
## Calculating cluster type_3
## Calculating cluster type_4
MD_DEGs2 <- MD_DEGs2 %>% arrange(desc(avg_log2FC))
write.xlsx(MD_DEGs2, file = here("jk_code", "DEGs_MD_SUBSUBclass.xlsx"), rowNames = TRUE)
# save as excel file
MD_DEGs2 %>%
group_by(cluster) %>%
dplyr::filter(avg_log2FC > 1, p_val_adj < 0.05) %>%
slice_head(n = 10) %>%
ungroup() -> top10
# save as excel file
DoHeatmap(SO4, features = top10$gene) + NoLegend()## Warning in DoHeatmap(SO4, features = top10$gene): The following features were
## omitted as they were not found in the scale.data slot for the SCT assay: Oasl2,
## Atp6v0d2, Irx3
## Warning: The `slot` argument of `FetchData()` is deprecated as of SeuratObject 5.0.0.
## ℹ Please use the `layer` argument instead.
## ℹ The deprecated feature was likely used in the Seurat package.
## Please report the issue at <https://github.com/satijalab/seurat/issues>.
## This warning is displayed once every 8 hours.
## Call `lifecycle::last_lifecycle_warnings()` to see where this warning was
## generated.